KSHV K-RTA interacts with OTUD4.

(A) Affinity purification followed by mass spectrometry analysis to identify K-RTA binding proteins. The number of identified peptides corresponding to K-RTA, NCOA2, and OTUD4 was summarized. (B) BCBL1-Tet-K-RTA cells were induced with Dox (1 μg/ml) and sodium butyrate (0.5 mM) for 48 h, and co-immunoprecipitation and immunoblotting were performed with the indicated antibodies. (C-E) HEK293T cells were transfected with the indicated plasmids, and WCLs were collected for immunoprecipitation with anti-FLAG affinity agarose, followed by immunoblotting. The interaction between K-RTA truncations, including K-RTAΔ490–535, K-RTAΔ536–589, K-RTAΔ590–650, and K-RTAΔ651–691, and OTUD4 was assessed by co-immunoprecipitation in HEK293T cells (C). The interaction between K-RTA truncations, including K-RTA (1–650), K-RTA (1–663), and K-RTA (1–676), and OTUD4 was assessed by co-immunoprecipitation (D). The interaction between K-RTA point mutations, including K-RTA(F689A), K-RTA(R690A), and K-RTA(D691A), and OTUD4 was assessed by co-immunoprecipitation (E).

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